Traditionally, platelets have been thought to promote the development of endovascular infections such as infective endocarditis (lE), by providing a adhesive surface upon damaged endothelium for colonization by circulating microorganisms. In contrast, recent evidence suggests that platelets serve an important host defense role against the development of endovascular infections at sites of endothelial damage via local secretion of endogenous microbicidal peptides , termed thrombin-induced platelet microbicidal protein or tPMP. Preliminary studies revealed the following data: i) tPMP kills the most common endovascular pathogens in nM concentrations; ii) microbial strains from bacteremic patients without IE were significantly more susceptible in vitro to the microbicidal action of tPMP than strains from bacteremic patients with IE. This suggested that phenotypic tPMP- resistance provides the organism with a survival advantage as regards induction of IE; iii) tPMP is synergistic in combination with conventional antibiotics in the killing and growth.inhibition of S. aureus ; and iv) the bacterial cell membrane appears to be a primary target for tPMP-induced lethality.The overall purpose of the current proposal is to define the fundamental microbicidal mechanisms of tPMP against S. aureus, the most virulent and commonest overall cause of endovascular infections, by: i) quantifying tPMP binding to the staphylococcal membrane; ii) delineating the role of bacterial transmembrane potential in the staphylocidal actions tPMP; iii) identifying membrane assembly of tPMP and subsequent membrane permeabilization and pore formation by tPMP; and iv) defining the mechanisms of phenotypic tPMP resistance, using genetically-related staphylococcal strains from a common genomic background which differ phenotypically in tPMP susceptibility. These studies may eventually define novel staphylocidal targets or unique staphylocidal mechanisms. These studies will also provide a solid foundation for defining the staphylocidal domains of tPMP by molecular biologic techniques. tPMP will also serve as a design template for development of synthetic congeners with potent antimicrobial activity.